Evolutionary sequence comparisons using high-density oligonucleotide arrays.
We explored the utility of high-density oligonucleotide arrays (DNA chips) for obtaining sequence information from homologous genes in closely related species. Orthologues of the human BRCA1 exon 11, all approximately 3.4 kb in length and ranging from 98.2% to 83.5% nucleotide identity, were subjected to hybridization-based and conventional dideoxysequencing analysis. Retrospective guidelines for identifying high-fidelity hybridization-based sequence calls were formulated based upon dideoxysequencing results. Prospective application of these rules yielded base-calling with at least 98.8% accuracy over orthologous sequence tracts shown to have approximately 99% identity. For higher primate sequences with greater than 97% nucleotide identity, base-calling was made with at least 99.91% accuracy covering a minimum of 97% of the sequence. Using a second-tier confirmatory hybridization chip strategy, shown in several cases to confirm the identity of predicted sequence changes, the complete sequence of the chimpanzee, gorilla and orangutan orthologues should be deducible solely through hybridization-based methodologies. Analysis of less highly conserved orthologues can still identify conserved nucleotide tracts of at least 15 nucleotides and can provide useful information for designing primers. DNA-chip based assays can be a valuable new technology for obtaining high-throughput cost-effective sequence information from related genomes.
Hacia
J G
JG
National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Makalowski
W
W
Edgemon
K
K
Erdos
M R
MR
Robbins
C M
CM
Fodor
S P
SP
Brody
L C
LC
Collins
F S
FS
D000508:Alouatta; D000818:Animals; D019313:BRCA1 Protein / Q000235:genetics*; D001483:Base Sequence; D017931:DNA Primers; D004285:Dogs; D019143:Evolution, Molecular; D005091:Exons; D005701:Galago; D019398:Genes, BRCA1; D005821:Genetic Techniques; D006071:Gorilla gorilla; D015186:Hominidae; D006801:Humans; D007900:Lemur; D008253:Macaca mulatta; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011148:Pongo pygmaeus; D011323:Primates / Q000145:classification / Q000235:genetics*
D019313:BRCA1 Protein; D017931:DNA Primers